Abstract:

Methods and compounds effective in ameliorating conditions characterized
by unwanted calcium channel activity, particularly unwanted N-type
calcium channel activity are disclosed. Specifically, a series of
compounds containing both a piperazine ring and a cyclopropyl ring are
disclosed of the general formula (I) where X1 and X2 are
linkers.
##STR00001##

33. The method of claim 30, wherein X1 is an optionally substituted
alkylene(1-2C).

34. The method of claim 33 wherein X1 is CH2 or CO.

35. The method of claim 30, wherein X2 is an optionally substituted
alkylene(1-2C).

36. The method of claim 35 wherein X2 is CH2 or CO.

37. The method of claim 30, wherein each Ar is independently an optionally
substituted phenyl ring.

38. The method of claim 37 wherein both Ar are unsubstituted phenyl.

39. The method of claim 30, wherein n is 0-1.

40. The method of claim 39, wherein n is 0.

41. The method of claim 30, wherein at least one R2 is other than H.

42. The method of claim 30, wherein the compound is a compound of formula
(2): ##STR00082## or a pharmaceutically acceptable salt or conjugate
thereof, wherein each A is independently H, or ═O;each Ar is
independently an optionally substituted phenyl ring; andeach R is
independently H, or an optionally substituted group selected from alkyl
(1-6C), alkenyl (2-6C), alkynyl (2-6C), heteroalkyl (2-6C), heteroalkenyl
(2-6C), heteroalkynyl (2-6C), aryl (6-10C), heteroaryl (5-12C),
C5-C12-heteroaryl-C1-C6-alkyl, and C6-C12-aryl-C1-C6-alkyl; and wherein
both R may together form an optionally substituted heterocyclic or
heteroaromatic ring.

46. The compound of claim 45 wherein X1 is an optionally substituted
alkylene(1-2C).

47. The compound of claim 45 wherein X1 is CH2 or CO.

48. The compound of claim 45, wherein X2 is an optionally substituted
alkylene(1-2C).

49. The compound of claim 48, wherein X2 is CH2 or CO.

50. The compound of claim 45, wherein each Ar is independently an
optionally substituted phenyl ring.

51. The compound of claim 50, wherein both Ar are unsubstituted phenyl.

52. The compound of claim 45, wherein n is 0-1.

53. The compound of claim 53 wherein n is 0.

54. The compound of claim 45, wherein at least one R2 is other than
H.

55. The compound of claim 45, which is a compound of formula (2):
##STR00084## or a pharmaceutically acceptable salt or conjugate thereof,
wherein each A is independently H, or ═O;each Ar is independently an
optionally substituted phenyl ring; andeach R is independently H, or an
optionally substituted group selected from alkyl (1-6C), alkenyl (2-6C),
alkynyl (2-6C), heteroalkyl (2-6C), heteroalkenyl (2-6C), heteroalkynyl
(2-6C), aryl (6-10C), heteroaryl (5-12C), C5-C12-heteroaryl-C1-C6-alkyl,
and C6-C12-aryl-C1-C6-alkyl; and wherein both R may together form an
optionally substituted heterocyclic or heteroaromatic ring.

58. A pharmaceutical composition which comprises a compound of claim 45,
in admixture with a pharmaceutically acceptable excipient.

Description:

TECHNICAL FIELD

[0001]The invention relates to compounds useful in treating conditions
associated with calcium channel function, and particularly conditions
associated with N-type calcium channel activity. More specifically, the
invention concerns compounds containing piperazine derivatives and also
possessing cyclopropyl functionality that are useful in treatment of
conditions such as stroke and pain.

[0003]Calcium channels have been shown to mediate the development and
maintenance of the neuronal sensitization processes associated with
neuropathic pain, and provide attractive targets for the development of
analgesic drugs (reviewed in Vanegas, H. & Schaible, H-G., Pain (2000)
85: 9-18). All of the high-threshold Ca channel types are expressed in
the spinal cord, and the contributions of L-, N and P/Q-types in acute
nociception are currently being investigated. In contrast, examination of
the functional roles of these channels in more chronic pain conditions
strongly indicates a pathophysiological role for the N-type channel
(reviewed in Vanegas & Schaible (2000) supra).

[0005]While its mechanism of action is not completely understood, current
evidence suggests that gabapentin does not directly interact with GABA
receptors in many neuronal systems, but rather modulates the activity of
high threshold calcium channels. Gabapentin has been shown to bind to the
calcium channel α2δ ancillary subunit, although it
remains to be determined whether this interaction accounts for its
therapeutic effects in neuropathic pain.

[0008]Ziconotide has been evaluated in a number of clinical trials via
intrathecal administration for the treatment of a variety of conditions
including post-herpetic neuralgia, phantom limb syndrome, HIV-related
neuropathic pain and intractable cancer pain (reviewed in Mathur, V. S.,
Seminars in Anesthesia. Perioperative Medicine and Pain (2000) 19:
67-75). In phase II and III clinical trials with patients unresponsive to
intrathecal opiates, ziconotide has significantly reduced pain scores and
in a number of specific instances resulted, in relief after many years of
continuous pain. Ziconotide is also being examined for the management of
severe post-operative pain as well as for brain damage following stroke
and severe head trauma (Heading. C., Curr Opin CPNS Investigasional Drugs
(1999) 1: 153-166). In two case studies ziconotide has been further
examined for usefulness in the management of intractable spasticity
following spinal cord injury in patients unresponsive to baclofen and
morphine (Ridgeway, B. et al., Pain (2000) 85: 287-289). In one instance,
ziconotide decreased the spasticity from the severe range to the mild to
none range with few side effects. In another patient, ziconotide also
reduced spasticity to the mild range although at the required dosage
significant side effects including memory loss, confusion and sedation
prevented continuation of the therapy.

[0010]U.S. Pat. No. 5,646,149 describes calcium channel antagonists of the
formula A-Y-B wherein B contains a piperazine or piperidine ring directly
linked to Y. An essential component of these molecules is represented by
A, which must be an antioxidant; the piperazine or piperidine itself is
said to be important. The exemplified compounds contain a benzhydryl
substituent, based on known calcium channel blockers (see below). U.S.
Pat. No. 5,703,071 discloses compounds said to be useful in treating
ischemic diseases. A mandatory portion of the molecule is a tropolone
residue, with substituents such as piperazine derivatives, including
their benzhydryl derivatives. U.S. Pat. No. 5,428,038 discloses compounds
indicated to exhibit a neural protective and antiallergic effect. These
compounds are coumarin derivativatives which may include derivatives of
piperazine and other six-membered heterocycles. A permitted substituent
on the heterocycle is diphenylhydroxymethyl. U.S. Pat. No. 6,458,781
describes 79 amides as calcium channel antagonists though only a couple
of which contain both piperazine rings and benzhydryl moieties. Thus,
approaches in the art for various indications which may involve calcium
channel blocking activity have employed compounds which incidentally
contain piperidine or piperazine moieties substituted with benzhydryl but
mandate additional substituents to maintain functionality.

[0011]Certain compounds containing both benzhydryl moieties and piperidine
or piperazine are known to be calcium channel antagonists and neuroleptic
drugs. For example, Gould, R. J., et al., Proc Natl Acad Sci USA (1983)
80:5122-5125 describes antischizophrenic neuroleptic drugs such as
lidoflazine, fluspirilene, pimozide, clopimozide, and penfluridol. It has
also been shown that fluspirilene binds to sites on L-type calcium
channels (King, V. K., et al., J Biol Chem (1989) 264:5633-5641) as well
as blocking N-type calcium current (Grantham, C. J., et al., Brit J
Pharmacol (1944) 111:483-488). In addition, Lomerizine, as developed by
Kanebo, K. K., is a known calcium channel blocker. However, Lomerizine is
not specific for N-type channels. A review of publications concerning
Lomerizine is found in Dooley, D., Current Opinion in CPNS
Investigational Drugs (1999) 1:116-125.

[0012]The present invention provides novel compounds having calcium
channel activity, and which are active as inhibitors of N-type calcium
channels in particular. These compounds are thus useful for treatment of
disorders including pain and certain mood disorders, gastrointestinal
disorders, genitourinary disorders, neurologic disorders and metabolic
disorders.

[0013]All patents, patent applications and publications identified herein
are hereby incorporated by reference in their entirety.

DISCLOSURE OF THE INVENTION

[0014]The invention relates to compounds useful in treating conditions
modulated by calcium channel activity and in particular conditions
mediated by N-type channel activity. The compounds of the invention are
heterocyclic compounds with structural features that enhance the calcium
channel blocking activity of the compounds. Thus, in one aspect, the
invention is directed to a method of treating conditions mediated by
calcium channel activity by administering to patients in need of such
treatment at least one compound of formula (1):

[0028]The invention is also directed to compounds of formula (1) or (2)
useful to modulate calcium channel activity, particularly N-type channel
activity, wherein the definition of such compound is as above. The
invention is also directed to the use of these compounds for the
preparation of medicaments for the treatment of conditions requiring
modulation of calcium channel activity, and in particular N-type calcium
channel activity. In another aspect, the invention is directed to
pharmaceutical compositions containing these compounds and to the use of
these compositions for treating conditions requiring modulation of
calcium channel activity, and particularly N-type calcium channel
activity.

DETAILED DESCRIPTION

[0029]As used herein, the term "alkyl," "alkenyl" and "alkynyl" include
straight-chain, branched-chain and cyclic monovalent substituents, as
well as combinations of these, containing only C and H when
unsubstituted. Examples include methyl, ethyl, isobutyl, cyclohexyl,
cyclopentylethyl, 2-propenyl, 3-butynyl, and the like. Typically, the
alkyl, alkenyl and alkynyl groups contain 1-8C (alkyl) or 2-8C (alkenyl
or alkynyl). In some embodiments, they contain 1-6C, 1-4C or 1-2C
(alkyl); or 2-6C or 2-4C (alkenyl or alkynyl). Further, any hydrogen atom
on one of these groups can be replaced with a halogen atom, and in
particular a fluoro or chloro, and still be within the scope of the
definition of alkyl, alkenyl and alkynyl. For example, CF3 is a 1C
alkyl. These groups may be also be substituted by other substituents.

[0030]Heteroalkyl, heteroalkenyl and heteroalkynyl are similarly defined
and contain at least one carbon atom but also contain one or more O, S or
N heteroatoms or combinations thereof within the backbone residue whereby
each heteroatom in the heteroalkyl, heteroalkenyl or heteroalkynyl group
replaces one carbon atom of the alkyl, alkenyl or alkynyl group to which
the heteroform corresponds. In preferred embodiments, the heteroalkyl,
heteroalkenyl and heteroalkynyl groups have C at each terminus to which
the group is attached to other groups, and the heteroatom(s) present are
not located at a terminal position. As is understood in the art, these
heteroforms do not contain more than three contiguous heteroatoms. In
preferred embodiments, the heteroatom is O or N. For greater certainty,
to the extent that alkyl is defined as 1-6C, then the corresponding
heteroalkyl contains 2-6 C, N, O, or S atoms such that the heteroalkyl
contains at least one C atom and at least one heteroatom. Similarly, when
alkyl is defined as 1-6C or 14C, the heteroform would be 2-6C or 2-4C
respectively, wherein one C is replaced by O, N or S. Accordingly, when
alkenyl or alkynyl is defined as 2-6C (or 2-4C), then the corresponding
heteroform would also contain 2-6 C, N, O, or S atoms (or 2-4) since the
heteroalkenyl or heteroalkynyl contains at least one carbon atom and at
least one heteroatom. Further, heteroalkyl, heteroalkenyl or
heteroalkynyl substituents may also contain one or more carbonyl groups.
Examples of heteroalkyl, heteroalkenyl and heteroalkynyl groups include
CH2OCH3, CH2 N(CH3)2, CH2OH,
(CH2)nNR2, OR, COOR, CONR2, (CH2)nOR,
(CH2)n, COR, (CH2)nCOOR, (CH2)nSR,
(CH2)nSOR, (CH2)nSO2R,
(CH2)nCONR2, NRCOR, NRCOOR, OCONR2, OCOR and the like
wherein the group contains at least one C and the size of the substituent
is consistent with the definition of alkyl, alkenyl and alkynyl.

[0031]As used herein, the terms "alkylene," "alkenyenyene" and
"alkynylene" refers to divalent groups having a specified size, typically
1-2C, 1-4C, 1-6C or 1-8C for the saturated groups and 2-4C, 2-6C or 2-8C
for the unsaturated groups. They include straight-chain, branched-chain
and cyclic forms as well as combinations of these, containing only C and
H when unsubstituted. Because they are divalent, they can link together
two parts of a molecule, as exemplified by X in formula (1). Examples
include methylene, ethylene, propylene, cyclopropan-1,1-diyl, ethylidene,
2-butene-1,4-diyl, and the like. These groups can be substituted by the
groups typically suitable as substituents for alkyl, alkenyl and alkynyl
groups as set forth herein. Thus C═O is a C1 alkylene that is
substituted by ═O, for example.

[0032]Heteroalkylene, heteroalkenylene and heteroalkynylene are similarly
defined as divalent groups having a specified size, typically 2-4C, 2-6C
or 2-8C for the saturated groups and 2-4C, 2-6C or 2-8C for the
unsaturated groups. They include straight chain, branched chain and
cyclic groups as well as combinations of these, and they further contain
at least one carbon atom but also contain one or more O, S or N
heteroatoms or combinations thereof within the backbone residue, whereby
each heteroatom in the heteroalkylene, heteroalkenylene or
heteroalkynylene group replaces one carbon atom of the alkylene,
alkenylene or alkynylene group to which the heteroform corresponds. As is
understood in the art, these heteroforms do not contain more than three
contiguous heteroatoms.

[0033]"Aromatic" moiety or "aryl" moiety refers to any monocyclic or fused
ring bicyclic system which has the characteristics of aromaticity in
terms of electron distribution throughout the ring system and includes a
monocyclic or fused bicyclic moiety such as phenyl or naphthyl;
"heteroaromatic" or "heteroaryl" also refers to such monocyclic or fused
bicyclic ring systems containing one or more heteroatoms selected from O,
S and N. The inclusion of a heteroatom permits inclusion of 5-membered
rings to be considered aromatic as well as 6-membered rings. Thus,
typical aromatic/heteroaromatic systems include pyridyl, pyrimidyl,
indolyl, benzimidazolyl, benzotriazolyl, isoquinolyl, quinolyl,
benzothiazolyl, benzofuranyl, thienyl, furyl, pyrrolyl, thiazolyl,
oxazolyl, imidazolyl and the like. Because tautomers are theoretically
possible, phthalimido is also considered aromatic. Typically, the ring
systems contain 5-12 ring member atoms or 6-10 ring member atoms. In some
embodiments, the aromatic or heteroaromatic moiety is a 6-membered
aromatic rings system optionally containing 1-2 nitrogen atoms. More
particularly, the moiety is an optionally substituted phenyl, 2-, 3- or
4-pyridyl, indolyl, 2- or 4-pyrimidyl, pyridazinyl, benzothiazolyl or
benzimidazolyl. Even more particularly, such moiety is phenyl, pyridyl,
or pyrimidyl and even more particularly, it is phenyl.

[0034]"O-aryl" or "O-heteroaryl" refers to aromatic or heteroaromatic
systems which are coupled to another residue through an oxygen atom. A
typical example of an O-aryl is phenoxy. Similarly, "arylalkyl" refers to
aromatic and heteroaromatic systems which are coupled to another residue
through a carbon chain, saturated or unsaturated, typically of 1-8C, 1-6C
or more particularly 1-4C when saturated or 2-8C, 2-6C or 2-4C when
unsaturated, including the heteroforms thereof. For greater certainty,
arylalkyl thus includes an aryl or heteroaryl group as defined above
connected to an alkyl, heteroalkyl, alkenyl heteroalkenyl, alkynyl or
heteroalkynyl moiety also as defined above. Typical arylalkyls would be
an aryl(6-12C)alkyl(1-8C), aryl(6-12C)alkenyl(2-8C), or
aryl(6-12C)alkynyl(2-8C), plus the heteroforms. A typical example is
phenylmethyl, commonly referred to as benzyl.

[0036]Optional substituents on a non-aromatic group, are typically
selected from the same list of substituents on aromatic or heteroaromatic
groups and may further be selected from ═O and ═NOR' where R' is
similarly defined. For greater certainty, two substituents on the same N
or adjacent C can form a 5-7 membered ring which may contain one or two
additional heteroatoms selected from N, O and S.

[0037]Halo may be any halogen atom, especially F, Cl, Br, or I, and more
particularly it is fluoro or chloro.

[0038]In general, any alkyl, alkenyl, alkynyl, or aryl (including all
heteroforms defined above) group contained in a substituent may itself
optionally be substituted by additional substituents. The nature of these
substituents is similar to those recited with regard to the substituents
on the basic structures above. Thus, where an embodiment of a substituent
is alkyl, this alkyl may optionally be substituted by the remaining
substituents listed as substituents where this makes chemical sense, and
where this does not undermine the size limit of alkyl per se; e.g., alkyl
substituted by alkyl or by alkenyl would simply extend the upper limit of
carbon atoms for these embodiments, and is not included. However, alkyl
substituted by aryl, amino, halo and the like would be included.

[0040]Ar is defined as an optionally substituted aromatic or
heteroaromatic ring. The two Ar groups may be the same or different; in
some embodiments they are the same. In certain embodiments each Ar
represent phenyl, so Ar2 CH-- represents a benzhydryl, and each
phenyl ring may independently be substituted or unsubstituted. In other
embodiments, Ar2 CH represents an unsubstituted benzhydryl.

[0041]X1 and XZ may independently be an optionally substituted
alkylene (1-4C) alkenylene (2-4C), alkynylene (2-4C), heteroalkylene
(2-4C), heteroalkenylene (2-4C), or heteroalkynylene (2-4C). In a more
particular embodiment, X1 and X2 may independently be an
optionally substituted 1-2C alkylene, and more particularly an optionally
substituted methylene. In even more particular embodiments. X1 and
X2 may independently be CH2 or C═O.

[0043]In some preferred embodiments, two or more of the particularly
described groups are combined into one compound: it is often suitable to
combine one of the specified embodiments of one feature as described
above with a specified embodiment or embodiments of one or more other
features as described above. For example, a specified embodiment includes
X1 as C═O, and another specified embodiment has both Ar as
optionally substituted phenyl groups (i.e., an optionally substituted
benzhydryl). Thus one preferred embodiment combines both of these
features together, i.e., X1 is CO in combination with both Ar
representing optionally substituted benzhydryl. In some specific
embodiments, n is 0 and in others n is 1. Thus additional preferred
embodiments include n=0 in combination with any of the preferred
combinations set forth above; other preferred combinations include n=1 in
combination with any of the preferred combinations set forth above.

[0044]The compounds of the invention may have ionizable groups so as to be
capable of preparation as salts. These salts may be acid addition salts
involving inorganic or organic acids or the salts may, in the case of
acidic forms of the compounds of the invention be prepared from inorganic
or organic bases. Frequently, the compounds are prepared or used as
pharmaceutically acceptable salts prepared as addition products of
pharmaceutically acceptable acids or bases. Suitable pharmaceutically
acceptable acids and bases are well-known in the art such as
hydrochloric, sulphuric, hydrobromic, acetic, lactic, citric, or tartaric
acids for forming acid addition salts, and potassium hydroxide, sodium
hydroxide, ammonium hydroxide, caffeine, various amines, and the like for
forming basic salts. Methods for preparation of the appropriate salts are
well-established in the art.

[0045]In some cases, the compounds of the invention contain one or more
chiral centers. The invention includes each of the isolated
stereoisomeric forms as well as mixtures of stereoisomers in varying
degrees of chiral purity, including racemic mixtures. It also encompasses
the various diastereomers and tautomers that can be formed. It expressly
includes both the cis and trans isomers of the cyclopropane rings shown
in Formula (1) and (2), although in some embodiments, the trans
cyclopropanes are preferred.

[0046]Compounds of formula (1) and (2) are also useful for the manufacture
of a medicament useful to treat conditions characterized by undesired
N-type calcium channel activities.

[0047]In addition, the compounds of the invention may be coupled through
conjugation to substances designed to alter the pharmacokinetics, for
targeting, or for other reasons. Thus, the invention further includes
conjugates of these compounds. For example, polyethylene glycol is often
coupled to substances to enhance half-life; the compounds may be coupled
to liposomes covalently or noncovalently or to other particulate
carriers. They may also be coupled to targeting agents such as antibodies
or peptidomimetics, often through linker moieties. Thus, the invention is
also directed to the compounds of formula (1) and (2) when modified so as
to be included in a conjugate of this type.

MODES OF CARRYING OUT THE INVENTION

[0048]The compounds of formula (1) and (2) are useful in the methods of
the invention and exert their desirable effects through their ability to
modulate the activity of calcium channels, particularly the activity of
N-type calcium channels. This makes them useful for treatment of certain
conditions where modulation of N-type calcium channels is desired,
including: chronic and acute pain; mood disorders such as anxiety,
depression, and addiction; neurodegenerative disorders; gastrointestinal
disorders such as inflammatory bowel disease and irritable bowel
syndrome; genitourinary disorders such as urinary incontinence,
interstitial colitis and sexual dysfunction; neuroprotection such as
cerebral ischemia, stroke and traumatic brain injury; and metabolic
disorders such as diabetes and obesity.

[0050]Anxiety as used herein includes but is not limited to the following
conditions: generalized anxiety disorder, social anxiety disorder, panic
disorder, obsessive-compulsive disorder, and post-traumatic stress
syndrome. Addiction includes but is not limited to dependence, withdrawal
and/or relapse of cocaine, opioid, alcohol and nicotine.

[0052]For greater certainty, in treating osteoarthritic pain, joint
mobility will also improve as the underlying chronic pain is reduced.
Thus, use of compounds of the present invention to treat osteoarthritic
pain inherently includes use of such compounds to improve joint mobility
in patients suffering from osteoarthritis.

[0053]It is known that calcium channel activity is involved in a
multiplicity of disorders, and particular types of channels are
associated with particular conditions. The association of N-type channels
in conditions associated with neural transmission would indicate that
compounds of the invention which target N-type receptors are most useful
in these conditions. Many of the members of the genus of compounds of
formula (1) and (2) exhibit high affinity for N-type channels. Thus, as
described below, they are screened for their ability to interact with
N-type channels as an initial indication of desirable function. It is
particularly desirable that the compounds exhibit IC50 values of
<1 μM. The IC50 is the concentration which inhibits 50% of the
calcium, barium or other permeant divalent cation flux at a particular
applied potential.

[0054]There are three distinguishable types of calcium channel inhibition.
The first, designated "open channel blockage," is conveniently
demonstrated when displayed calcium channels are maintained at an
artificially negative resting potential of about -100 mV (as
distinguished from the typical endogenous resting maintained potential of
about -70 mV). When the displayed channels are abruptly depolarized under
these conditions, calcium ions are caused to flow through the channel and
exhibit a peak current flow which then decays. Open channel blocking
inhibitors diminish the current exhibited at the peak flow and can also
accelerate the rate of current decay.

[0055]This type of inhibition is distinguished from a second type of
block, referred to herein as "inactivation inhibition." When maintained
at less negative resting potentials, such as the physiologically
important potential of -70 mV, a certain percentage of the channels may
undergo conformational change, rendering them incapable of being
activated--i.e., opened--by the abrupt depolarization. Thus, the peak
current due to calcium ion flow will be diminished not because the open
channel is blocked, but because some of the channels are unavailable for
opening (inactivated). "Inactivation" type inhibitors increase the
percentage of receptors that are in an inactivated state.

[0056]A third type of inhibition is designated "resting channel block".
Resting channel block is the inhibition of the channel that occurs in the
absence of membrane depolarization, that would normally lead to opening
or inactivation. For example, resting channel blockers would diminish the
peak current amplitude during the very first depolarization after drug
application without additional inhibition during the depolarization.

[0057]In order to be maximally useful in treatment, it is also helpful to
assess the side reactions which might occur. Thus, in addition to being
able to modulate a particular calcium channel, it is desirable that the
compound has very low activity with respect to the hERG K.sup.+ channel
which is expressed in the heart. Compounds that block this channel with
high potency may cause reactions which are fatal. Thus, for a compound
that modulates the calcium channel, it should also be shown that the hERG
K.sup.+ channel is not inhibited. Similarly, it would be undesirable for
the compound to inhibit cytochrome p450 since this enzyme is required for
drug detoxification. Finally, the compound will be evaluated for calcium
ion channel type specificity by comparing its activity among the various
types of calcium channels, and specificity for one particular channel
type is preferred. The compounds which progress through these tests
successfully are then examined in animal models as actual drug
candidates.

[0058]The compounds of the invention modulate the activity of calcium
channels; in general, said modulation is the inhibition of the ability of
the channel to transport calcium. As described below, the effect of a
particular compound on calcium channel activity can readily be
ascertained in a routine assay whereby the conditions are arranged so
that the channel is activated, and the effect of the compound on this
activation (either positive or negative) is assessed. Typical assays are
described hereinbelow in Examples 3 and 4.

[0059]Libraries and Screening

[0060]The compounds of the invention can be synthesized individually using
methods known in the art per se, or as members of a combinatorial
library.

[0061]Synthesis of combinatorial libraries is now commonplace in the art.
Suitable descriptions of such syntheses are found, for example, in
Wentworth, Jr., P., et al., Current Opinion in Biol. (1993) 9:109-115;
Salemme, F. R., et al., Structure (1997) 5:319-324. The libraries contain
compounds with various substituents and various degrees of unsaturation,
as well as different chain lengths. The libraries, which contain, as few
as 10, but typically several hundred members to several thousand members,
may then be screened for compounds which are particularly effective
against a specific subtype of calcium channel, e.g., the N-type channel.
In addition, using standard screening protocols, the libraries may be
screened for compounds that block additional channels or receptors such
as sodium channels, potassium channels and the like.

[0062]Methods of performing these screening functions are well known in
the art. These methods can also be used for individually ascertaining the
ability of a compound to agonize or antagonize the channel. Typically,
the channel to be targeted is expressed at the surface of a recombinant
host cell such as human embryonic kidney cells. The ability of the
members of the library to bind the channel to be tested is measured, for
example, by the ability of the compound in the library to displace a
labeled binding ligand such as the ligand normally associated with the
channel or an antibody to the channel. More typically, ability to
antagonize the channel is measured in the presence of calcium, barium or
other permeant divalent cation and the ability of the compound to
interfere with the signal generated is measured using standard
techniques. In more detail, one method involves the binding of
radiolabeled agents that interact with the calcium channel and subsequent
analysis of equilibrium binding measurements including, but not limited
to, on rates, off rates, Kd values and competitive binding by other
molecules.

[0063]Another method involves the screening for the effects of compounds
by electrophysiological assay whereby individual cells are impaled with a
microelectrode and currents through the calcium channel are recorded
before and after application of the compound of interest.

[0064]Another method, high-throughput spectrophotometric assay, utilizes
loading of the cell lines with a fluorescent dye sensitive to
intracellular calcium concentration and subsequent examination of the
effects of compounds on the ability of depolarization by potassium
chloride or other means to alter intracellular calcium levels.

[0065]As described above, a more definitive assay can be used to
distinguish inhibitors of calcium flow which operate as open channel
blockers, as opposed to those that operate by promoting inactivation of
the channel or as resting channel blockers. The methods to distinguish
these types of inhibition are more particularly described in the examples
below. In general, open-channel blockers are assessed by measuring the
level of peak current when depolarization is imposed on a background
resting potential of about -100 mV in the presence and absence of the
candidate compound. Successful open-channel blockers will reduce the peak
current observed and may accelerate the decay of this current. Compounds
that are inactivated channel blockers are generally determined by their
ability to shift the voltage dependence of inactivation towards more
negative potentials. This is also reflected in their ability to reduce
peak currents at more depolarized holding potentials (e.g., -70 mV) and
at higher frequencies of stimulation, e.g., 0.2 Hz vs. 0.03 Hz. Finally,
resting channel blockers would diminish the peak current amplitude during
the very first depolarization after drug application without additional
inhibition during the depolarization.

[0066]Accordingly, a library of compounds of formula (1) or formula (2)
can be used to identify a compound having a desired combination of
activities that includes activity against at least one type of calcium
channel. For example, the library can be used to identify a compound
having a suitable level of activity on N-type calcium channels while
having minimal activity on HERG K+ channels.

[0067]Utility and Administration

[0068]For use as treatment of human and animal subjects, the compounds of
the invention can be formulated as pharmaceutical or veterinary
compositions. Depending on the subject to be treated, the mode of
administration, and the type of treatment desired--e.g., prevention,
prophylaxis, therapy; the compounds are formulated in ways consonant with
these parameters. A summary of such techniques is found in Remington's
Pharmaceutical Sciences, latest edition, Mack Publishing Co., Easton,
Pa., incorporated herein by reference.

[0069]In general, for use in treatment, the compounds of formula (1) or
(2) may be used alone, as mixtures of two or more compounds of formula
(1) and/or (2) or in combination with other pharmaceuticals. An example
of other potential pharmaceuticals to combine with the compounds of
formula (1) and (2) would include pharmaceuticals for the treatment of
the same indication but having a different mechanism of action from
N-type calcium channel blocking. For example, in the treatment of pain, a
compound of formula (1) or (2) may be combined with another pain relief
treatment such as an NSAID, or a compound which selectively inhibits
COX-2, or an opioid, or an adjuvant analgesic such as an antidepressant.
Another example of a potential pharmaceutical to combine with the
compounds of formula (1) or (2) would include pharmaceuticals for the
treatment of different yet associated or related symptoms or indications.
Depending on the mode of administration, the compounds will be formulated
into suitable compositions to permit facile delivery.

[0070]The compounds of the invention may be prepared and used as
pharmaceutical compositions comprising an effective amount of at least
one compound of formula (1) or (2) admixed with a pharmaceutically
acceptable carrier or excipient, as is well known in the art.
Formulations may be prepared in a manner suitable for systemic
administration or topical or local administration. Systemic formulations
include those designed for injection (e.g., intramuscular, intravenous or
subcutaneous injection) or may be prepared for transdermal, transmucosal,
or oral administration. The formulation will generally include a diluent
as well as, in some cases, adjuvants, buffers, preservatives and the
like. The compounds can be administered also in liposomal compositions or
as microemulsions.

[0071]For injection, formulations can be prepared in conventional forms as
liquid solutions or suspensions or as solid forms suitable for solution
or suspension in liquid prior to injection or as emulsions. Suitable
excipients include, for example, water, saline, dextrose, glycerol and
the like. Such compositions may also contain amounts of nontoxic
auxiliary substances such as wetting or emulsifying agents, pH buffering
agents and the like, such as, for example, sodium acetate, sorbitan
monolaurate, and so forth.

[0072]Various sustained release systems for drugs have also been devised.
See, for example, U.S. Pat. No. 5,624,677.

[0073]Systemic administration may also include relatively noninvasive
methods such as the use of suppositories, transdermal patches,
transmucosal delivery and intranasal administration. Oral administration
is also suitable for compounds of the invention. Suitable forms include
syrups, capsules, tablets, as is understood in the art.

[0074]For administration to animal or human subjects, the dosage of the
compounds of the invention is typically 0.01-15 mg/kg, preferably 0.1-10
mg/kg. However, dosage levels are highly dependent on the nature of the
condition, drug efficacy, the condition of the patient, the judgment of
the practitioner, and the frequency and mode of administration.
Optimization of the dosage for a particular subject is within the
ordinary level of skill in the art.

[0075]Synthesis of the Invention Compounds

[0076]The following reaction schemes and examples are intended to
illustrate the synthesis of a representative number of compounds.
Accordingly, the following examples are intended to illustrate but not to
limit the invention. Additional compounds not specifically exemplified
may be synthesized using conventional methods in combination with the
methods described hereinbelow.

[0079]Diethyl (1R,2R)-1,2-cyclopropanedicarboxylate 3.5 ml (20 mmol) and
NaOH (0.8 g, 20.4 mmol) in EtOH (60 ml) and water (5 ml) was stirred at
room temperature overnight. The mixture was then concentrated to remove
the solvent. The residue was redissolved in water and washed with diethyl
ether (40 ml) to remove any unreacted starting material. The aqueous
solution was then adjusted to pH˜2 with 2N HCl and extracted with
ethyl acetate (2×40 ml). The combined organic extracts were dried
over anhydrous sodium sulfate and concentrated to give crude monoester
monoacid (2.8 g, 90%).

[0090]Oxalyl chloride (0.250 ml, 2.87 mmol) in 5 ml of DCM was cooled to
-78° C., followed by the addition of dimethyl sulfoxide (DMSO,
0.465 ml, pre-solvated in 1 ml of DCM). The solutions were stirred at
-78° C. for about 15 min.
(1R,2R)-(4-benzhydrylpiperazin-1-yl)(2-(hydroxymethyl)cyclopropyl)methano-
ne (0.87 g, 2.5 mmol) in DCM (5 ml) was then added and the reaction
mixture was stirred further for 30 min. triethylamine (TEA, 1.75 ml) was
then added and the reaction mixture was allowed to come to room
temperature with stirring continued for further 30 min. Water (3 ml) was
finally added to quench the reaction and the organic layer was separated,
dried over anhydrous sodium sulfate and concentrated to yield the
(1R,2R)-2-(4-benzhydrylpiperazin-1-carbonyl)cyclopropanecarbaldehyde (0.7
g crude, 80%).

[0096]Succinic anhydride (10 g), (1R,2S,5R)-(-) menthol (32 g), 200 mg of
p-toluenesulfonic acid and 100 ml of toluene were refluxed overnight
using a Dean Stark trap. The mixture was cooled and diluted with 100 ml
hexane and poured into 15 ml of saturated sodium bicarbonate, methanol
(70 ml) and water (100 ml). The organic phase was separated and the
aqueous phase was extracted with hexane (2×) and the combined
organic layers were washed with brine and dried. 50 ml methanol was added
and left in the refrigerator overnight. The following day, the
precipitate was collected to give 30 g of
(1R,2S,5R)-2-isopropyl-5-methylcyclohexyl)
(1S,2S,5R)-2-isopropyl-5-methylcyclohexyl succinate.

[0098]To a cooled solution of 2,2,6,6-tetramethylpiperidine (15 ml) in dry
THF (100 ml), 40 ml of n-butyllithium was added dropwise over 10 minutes.
The mixture was then cooled to -78° C. followed by addition of
(1R,2S,5R)-2-isopropyl-5-methylcyclohexyl)
(1S,2S,5R)-2-isopropyl-5-methylcyclohexyl succinate (1.5 g) over 10 min
and then 2.2 ml of bromochloromethane was added dropwise over a period of
10 min. The mixture was stirred for two hours and then 1.5 ml of
isobutyraldehyde was added to quench the unreacted anion and the reaction
mixture was then stirred for an additional 30 min. The reaction mixture
was poured onto 1N HCl and the reaction product was extracted with ether
(3×100 ml), washed with brine and then dried. Column chromatography
(18:1-hexane-ether) was used to give of
(1R,2S,5R)-2-isopropyl-5-methylcyclohexyl)
2-(1S,2S,5R)-2-isopropyl-5-methylcyclohexyl)-cyclopropane-1,2-dicarboxyla-
te in 85% yield.

[0100](1R,2S,5R)-2-isopropyl-5-methylcyclohexyl)
2-(1S,2S,5R)-2-isopropyl-5-methylcyclohexyl)cyclopropane-1,2-dicarboxylat-
e (1.5 g), NaOH (160 mg), water (0.8 ml), isopropyl alcohol (10 ml) were
combined and stirred at 50° C. overnight and then stirred at
85° C. for 2 hr. The reaction mixture was then concentrated and
the reaction product was extracted with ether and 5 ml water. The aqueous
layer was acidified with 2 N HCl and extracted with ether (3×40)
and dried to give the desired product in 90% yield.

[0102]A solution of
(1S,2S)-2-(((1S,2S,5R)-2-isopropyl-5-methylcyclohexyloxy)
carbonyl)cyclopropanecarboxylic acid (16 g), diphenylmethyl piperazine
(0.5 g), EDC (1.7 g) and DMAP (trace) in dichloromethane (20 ml) was
stirred at room temperature overnight, and then concentrated. Water was
added to the reaction mixture and the reaction product was then extracted
with ethyl acetate (2×25 ml). The combined organic solution was
dried over sodium sulfate and concentrated. The residue was applied to
flash column chromatography using petroleum ether and ethyl acetate (1:1)
as eluents to give
(1S,2S)-((1S,2S,5R)-2-isopropyl-5-methylcyclohexyl)-2-((4-benzhydrylpiper-
azin-1-carbonyl)cyclopropanecarboxylate in 76% yield.

[0104]A solution of
(1S,2S)-((1S,2S,5R)-2-isopropyl-5-methylcyclohexyl)2-((4-benzhydrylpipera-
zin-1-carbonyl)cyclopropanecarboxylate (2.2 g), LiOH--H2O (1 g), THE
(15 ml) was heated at 60° C. for 5 hr. The reaction mixture was
then evaporated. Water (10 ml) was added to the reaction mixture and the
reaction product was extracted with ether (2×20 ml) and then the
aqueous phase was acidified with 6N HCl to pH 4.5 and the precipitate was
filtered and dried in vacuum to give the desired carboxylic acid in 90%
yield.

[0109]Ethyl 2-formyl-1-cyclopropanecarboxylate (4.5 g),
diphenylmethylpiperazin (8.5 g) and triacetoxysodium borohydride (9.5) g
in dichloroethane (100 ml) was stirred overnight. The solvent was then
evaporated and the residue was dissolved in ethyl acetate (100 ml)
followed by washing with sodium bicarbonate. The organic layer was dried
over sodium sulfate and evaporated. The residue was applied to column
chromatography using ethyl acetate as eluents to give ethyl
2-{[4-(diphenylmethyl)piperazin 1-yl]methyl}cyclopropanecarboxylate in
92% yield.

[0111]A solution of ethyl
2-{[4-(diphenylmethyl)piperazin-1-yl]methyl}-cyclopropanecarboxylate (12
g), LiOH--H20 (10 g), water (30 ml), THF (100 ml), methanol (15 ml)
was stirred overnight at room temperature. The reaction mixture was
evaporated and the residue was then redissolved in water (40 ml) and
acidified with 6N HCl to pH 5. The aqueous layer was then extracted with
ethyl acetate to give 10 g of
2-{[4-(diphenylmethyl)piperazin-1-yl]methyl}cyclopropanecarboxylic acid
in 95% yield.

[0114]Following the general procedures set forth in Reaction Schemes 1-4
and Examples 1-15, the following compounds listed in Table 3 below were
prepared. Mass spectrometry was employed with the final compound and at
various stages throughout the synthesis as a confirmation of the identity
of the product obtained (M+1). For the mass spectrometric analysis,
samples were prepared at an approximate concentration of 1 μg/mL in
acetonitrile with 0.1% formic acid. Samples were then manually infused
into an Applied Biosystems API13000 triple quadrupole mass spectrometer
and scanned in Q1 in the range of 50 to 700 m/z.

[0120]The effects of intrathecally delivered compounds of the invention on
the rat formalin model can also be measured. The compounds can be
reconstituted to stock solutions of approximately 10 mg/ml in propylene
glycol. Typically eight Holtzman male rats of 275-375 g size are randomly
selected per test article.

[0121]The following study groups can be used, with test article, vehicle
control (propylene glycol) and saline delivered intraperitoneally (IP):

[0122]Prior to initiation of drug delivery baseline behavioral and testing
data can be taken. At selected times after infusion of the Test or
Control Article these data can then be again collected.

[0123]On the morning of testing, a small metal band (0.5 g) is loosely
placed around the right hind paw. The rat is placed in a cylindrical
Plexiglas chamber for adaptation a minimum of 30 minutes. Test Article or
Vehicle Control Article is administered 10 minutes prior to formalin
injection (50 μl of 5% formalin) into the dorsal surface of the right
hindpaw of the rat. The animal is then placed into the chamber of the
automated formalin apparatus where movement of the formalin injected paw
is monitored and the number of paw flinches tallied by minute over the
next 60 minutes (Malmberg, A. B., et al., Anesthesiology (1993)
79:270-281).

[0124]Results can be presented as Maximum Possible Effect±SEM, where
saline control=100%.

Example 19

Spinal Nerve Ligation Model of Neuropathic Pain

[0125]Spinal nerve ligation (SNL) injury can be induced using the
procedure of Kim and Chung, (Kim, S. H., et al., Pain (1992) 50:355-363)
in male Sprague-Dawley rats (Harlan; Indianapolis, Ind.) weighing 200 to
300 grams. Anesthesia is induced with 2% halothane in O2 at 2 L/min
and maintained with 0.5% halothane in O2. After surgical preparation
of the rats and exposure of the dorsal vertebral column from L4 to
S2, the L5 and L6 spinal nerves are tightly ligated distal
to the dorsal root ganglion using 4-0 silk suture. The incision is
closed, and the animals are allowed to recover for 5 days. Rats that
exhibit motor deficiency (such as paw-dragging) or failure to exhibit
subsequent tactile allodynia are excluded from further testing. Sham
control rats undergo the same operation and handling as the experimental
animals, but without SNL.

[0126]The assessment of tactile allodynia consists of measuring the
withdrawal threshold of the paw ipsilateral to the site of nerve injury
in response to probing with a series of calibrated von Frey filaments.
Each filament is applied perpendicularly to the plantar surface of the
ligated paw of rats kept in suspended wire-mesh cages. Measurements are
taken before and after administration of drug or vehicle. Withdrawal
threshold is determined by sequentially increasing and decreasing the
stimulus strength ("up and down" method), analyzed using a Dixon
non-parametric test (Chaplan S. R., et al., J Pharmacol Exp Ther (1994)
269:1117-1123), and expressed as the mean withdrawal threshold.

[0127]The method of Hargreaves and colleagues (Hargreaves, K., et al, Pain
(1988) 32:77-8) can be employed to assess paw-withdrawal latency to a
thermal nociceptive stimulus. Rats are allowed to acclimate within a
plexiglas enclosure on a clear glass plate maintained at 3° C. A
radiant heat source (i.e., high intensity projector lamp) is then
activated with a timer and focused onto the plantar surface of the
affected paw of nerve-injured or carrageenan-injected rats.
Paw-withdrawal latency can be determined by a photocell that halted both
lamp and timer when the paw is withdrawn. The latency to withdrawal of
the paw from the radiant heat source is determined prior to carrageenan
or L5/L5 SNL, 3 hours after carrageenan or 7 days after L5/L6 SNL but
before drug and after drug administration. A maximal cut-off of 40
seconds is employed to prevent tissue damage. Paw withdrawal latencies
can be thus determined to the nearest 0.1 second. Reversal of thermal
hyperalgesia is indicated by a return of the paw withdrawal latencies to
the pre-treatment baseline latencies (i.e., 21 seconds). Anti-nociception
is indicated by a significant (p<0.05) increase in paw withdrawal
latency above this baseline. Data is converted to % anti hyperalgesia or
% anti-nociception by the formula: (100×(test latency-baseline
latency)/(cut-off-baseline latency) where cut-off is 21 seconds for
determining anti-hyperalgesia and 40 seconds for determining
anti-nociception.